Microtome knife clamp with knife cooling means



; NOV. 30, 1965 s oN MICROTOME KNIFE CLAMP WITH KNIFE COOLING MEANS 3Sheets-Shet 1 Filed Oct. 1, 1963 /N (ISA/Tot Nov. 30, 1965 E. R. SHANDON3,220,290

MICROTOME KNIFE CLAMP WITH KNIFE COOLING MEANS Filed Oct. 1, 1963 3Sheets-Sheet 2 560 57 521227 Saw/v00 Nov. 30, 1965 E. R. SHANDON3,220,290

MIGROTOME KNIFE CLAMP WITH KNIFE COOLING MEANS Filed Oct. 1, 1963 3Sheets-Sheet. 3

United States Patent 3,220,290 MICROTOME KNIFE CLAMP WITH KNIFE COOLINGMEANS Ernest Robert Shandon, London, England, assignor, by mesneassignments, to C. Reichert Optische Werke A.G., Vienna, Austria, anAustrian company Filed Oct. 1, 1963, Ser. No. 312,957 Claims priority,application Great Britain, Oct. 15, 1962, 38,957/ 62 5 Claims. (Cl.83-171) This invention relates to microtomes and, more specif ically, toa microtome suitable for cutting sections from a frozen specimen.

The microtome is a precision instrument for enabling Wafer-thin sectionsof accurately controlled thickness, which may be of the order of a fewmicrons, to be taken from specimens mounted thereon and comprisesessentially a massive stand having a clamp holder for clamping inposition an object clamp in which the specimen can be held, and a knifeclamp in which a replaceable microtome knife may be gripped. To take asection from the block the two clamps are moved relative to one anotherso that the cutting edge of the knife slices across the face of theblock or specimen at a correct inclination and tilt to produce a sectionof the desired thickness.

The texture of some specimens is so soft that if attempts are made tosection them directly using the microtome the sections produced are soseverely distorted or damaged as to be useless. To handle suchspecimens, the microtome is commonly provided with a freezing stagewhich can be substituted for the object clamp and is equipped with meansfor freezing the specimen. Once the specimen is frozen it adheres firmlyto the freezing stage and its texture is made more suitable forwithstanding sectioning. Freezing is conveniently effected by passingover the specimen a stream of carbon dioxide.

To take sections, the microtome knife must also be reduced to thetemperature of the specimen as otherwise the relative warmth of theknife would melt the section whilst being cut so that it collapses intoa wet slime on the knife. One method of suitably cooling the knife is toplay on its surface, prior to the out being made, a jet of coolingcarbon dioxide obtained from the same source as is used to freeze thespecimen.

A second method which has been proposed is to clamp a thermo-electriccooling device, such as a Peltier cooling junction, directly onto thesurface of the microtome knife and then to cool the knife by connectingthe device to a suitable electrical circuit. By passing a unidirectionalelectric current through the circuit in the correct direction, heat willbe extracted from the microtome knife by the cooling device andliberated at the heat sink which can be located remote from the knife.However, as microtome knives require frequent replacement they musteither be individually provided with cooling devices which would be bothexpensive and cumbersome, or the cooling device must be so arranged thatit can be transferred from knife to knife between the fitting ofsuccessive knives to the microtome. Such a process would be laborious tocarry out. Furthermore, it is not practicable to use this method ofelectrical cooling on microtome knives less than centimetres in lengthas there is inadequate space on the knife to mount the cooling device.

ice

An object of this invention is the provision of improved means forcooling a microtome knife.

Another object of the invention is to enable a standard, unmodifiedmicrotome knife to be cooled for slicing frozen sections without havingintermittently to play a stream of cooling fluid on its cutting edge.

A further object of the invention is to provide a microtome with theadvantage obtained by using electrical cooling equipment for cooling amicrotome knife and the advantage obtained by being able to use astandard, unmodified microtome knife with the microtome.

An additional object of the invention is the provision of anelectrically cooled microtome knife clamp having means for ensuring thatthe knife temperature remains below a selected temperature.

Yet another object of the invention is the provision of a microtome withdual purpose clamping means serving both to grip the knife and toprovide part of a thermally conductive path extending from the knife toa heat sink inside the knife clamp.

In accordance with the present invention, a microtome knife clamp isprovided with jaws for gripping a knife, a mechanism for opening andclosing the jaws to release and to hold the knife respectively, a heatsink provided with a cooling unit built into the clamp, and aheat-transmitting element terminating in a pad for contacting the knifeand extending to the heat sink.

A microtome knife is kept cold when gripped by a knife clamp constructedin accordance with the invention, by

the conduction of heat from the knife by way of the padand heattransmitting element to the heat sink which is maintained cool byoperation of the cooling unit. The

knife clamp of the invention is able to receive standard,

unmodified microtome knives and thus sectioning of frozen specimens canbe effected quickly and easily without having to attach to individualmicrotome knives spe-- cial cooling apparatus and without having toprovide special cooling apparatus for cooling the knife by discharging astream of cooling fiuid onto it.

The cooling unit on the knife clamp may comprise a reservoir throughwhich cooling carbon dioxide or other coolant fluid can be passed.Alternatively, the reservoir may be adapted to receive a charge ofliquid air or other cooling fluid which extracts heat from the knifeduring vaporisation of the fluid. Suitably, the heat conducting elementcomprises a plate of material having good heat conducting propertiessuch as silver, aluminum or copper and having one side forming one wallof the reservoir or heat sink and the other side shaped to form a padhaving a surface contour complementary to that of the surface of themicrotome knife which is to be engaged by the pad.

Means may also be provided on the knife clamp for maintaining the knifebelow a desired maximum temperature and/or for measuring itstemperature. Such means may comprise a thermistor arranged to control anelectrical circuit which regulates the operation of the cooling unit.Alternatively, or additionally, the thermistor may be arranged tocontrol a suit-able circuit connected to temperature indicating meanswhich provide an indication of the knife temperature.

The preferred form of cooling unit, however, comprises a thermo-electriccooling device or devices connectable in an electric circuit energisableto cause the cooling device to extract heat from its immediate vicinity.One

suitable form of thermo-electric cooling device is a Peltier' junctionwhich is cooled when a unidirectional electrical current is passedthrough it in a suitable direction. A Peltier heating junction isconnected in series with the cooling junction and arranged at somedistance from the knife at one side of a cooling chamber through which acooling fluid is passed. The electrical current flowing through bothPeltier junctions causes the cooling junction to extract heat which isliberated at the other junction and is removed from the vicinity of theclamp by the cooling fluid.

The knife clamp of the invention may be used separately from a microtomeor may be fixed permanently to it as is usually the case withprecision-built microtomes. Thus the clamp of the invention is suitablycombined with a microtome having a massive stand, a holder for afreezing stage, and means for effective relative movement between theclamp and the holder so that a section can be sliced from a frozenspecimen on the freezing stage.

Suitably the microtome is also provided with an object clamp which maybe mounted in the holder in place of the freezing stage when it isdesired to take sections from an object block or a specimen notrequiring freezing. The invention will now be described in more detail,by way of examples, with reference to the accompanying drawings, inwhich:

FIGURE 1 is a diagrammatic perspective view of the principal parts of amicrotome;

' FIGURE 2 is a plan view of a two-part knife clamp used on themicrotome;

FIGURE 3 is a section through part of the knife clamp shown in FIGURE 2and taken on the line indicated by the arrows IIIIII in that figure;

FIGURE 4 shows in plan a part of a knife clamp of similar form to thatshown in FIGURE 2, but modified in accordance with the second embodimentof the invention; and

FIGURE 5 is a section through FIGURE 4 taken on the line indicated bythe arrows V-V in that figure.

FIGURE 1 shows a microtome comprising a massive stand 1 adapted to beclamped to an edge of a table (not shown) and provided with an uprightlimb 2 Serving to hold in place a vertical spindle 3 which extendsalongside the limb 2 and is pivoted at its ends thereto. A horizontalcarrier arm 4 is fixed at one end to the upper end of the spindle 3 andcarries at its free end a knob 5 which is gripped by the hand duringarcuate movements of the arm 4 about the spindle axis.

A box-like knife clamp 6 is rotatably mounted on the carrier arm 4 andcomprises a hinged, split two-part casting, the two parts of which areadapted to be drawn together by two screws 11 and respectively compriseupper and lower jaws 7 and 8 for gripping the marginal portions of theupper and lower surfaces of a knife 12 well away from the centralportion of its cutting edge 13. Each jaw is provided at each end with aprojection 14 extending laterally away from the carrier arm 4 and forengaging a surface of the knife at one end thereof.

Referring to FIGURES 2 and 3 each projection 14 is provided adjacent theposition of the gripped knife 12 with a heat sink which includes acooling unit built into the clamp. The heat sink includes a metal plate23 which is shaped on one side to form a pad of complementary shape tothe surface of the knife 12 and to contact such surface when the knifeis gripped, as shown, between the jaws 7 and 8. The plate 23 is made ofa metal having good heat-conducting properties, such as silver, copperor aluminum, so as to provide a good heat-conducting path betwen theknife 12 and a set of Peltier cooling junctions 15 bonded to the otherside of the plate 23.

The Peltiervcooling junctions 15 are connected by a number ofsemiconductor elements 20 to a set of Peltier heating junctions 17. Theheating junctions 17 are bonded to one face of a plate 123 which is alsomade of a good heat-conducting metal and has its other face located atone side of a chamber 18 formed inside the projection 14. The chamber 18is provided at opposite ends with an inlet 21 and an outlet 22 forpassing cooling water through the chamber and the two chambers 18 ateach end of the knife 12 are interconnected by a piece of flexibletubing 28 for serial flow of water between them. The tubing 28 allowsthe jaws to be opened and closed without interference when the microtomeknife requires replacing with a sharper knife.

Each jaw of the knife clamp is additionally provided with a temperaturesensing device comprising a thermistor 24 mounted alongside one of thePeltier cooling junctions 15. Electrical terminals 27 on each jaw enableexternal electrical circuiting to be connected to the two sets ofjunctions 15 and 17 and to the thermistor 24, so that the electricalcurrent flowing through the junctions can be controlled in accordancewith the temperature as sensed by the thermistor 24. The thermistor 24may be connected to a control circuit which regulates the currentthrough the junctions 15 and 17 automatically, and/or it may beconnected to an indicator from which the knife temperature may be readdirectly by the microtome user. Each thermistor 24 may be provided withadjusting means (not shown) alterable to change the setting of thethermistor and thus enable the temperature of the knife to be closelyregulated and/ or measured.

All the surfaces of the jaws, apart from the metal plates 23 which arein contact with the gripped knife may be provided with a layer ofthermal insulation 30 to reduce the transfer of heat therebetween. Thelayer of insulation may comprise a plastic material. As will be notedfrom FIGURE 2, the screws 11 for drawing the two jaws together are sopositioned that they do not come in contact with the microtome knifewhen it is gripped between the jaws so that heat conduction to the knifefrom the screws is prevented.

The knife clamp is provided with two rearwardly extending clampingscrews 31 which enable it to be fixed in a desired angular position onthe carrier arm 4. By slackening these screws the angle of attack of theknife to the specimen to be sectioned can be altered.

Referring again to FIGURE 1, the path of movement of the knife isdetermined by an arcuate track 42 coaxially arranged around the axis ofthe spindle 3 along the top of a curved web 43. The end portion of thecarrier arm 4 adjacent the knob 5 is provided with a pad (not shown)which is guided by the track 42 during cutting movement of the knifeacross the top of the specimen. The engagement of the pad and the trackensures that any vertical forces applied to the knob 5 do not influencethe thickness of the section cut from the specimen. A spring return (notshown) normally biases the carrier arm to some intermediate position onthe track.

The specimen 36 to be sectioned is mounted on a freezing stage 32 whichis removably mounted in an object clamp holder 34 and held therein by alocking device 35. The freezing stage 32 may be removed and replaced byan object clamp (not shown) when desired, when it is required to sectionan object block which does not have to be frozen.

The freezing stage 32 shown in FIGURE 1 comprises a drum having anannularly grooved upper surface to which water can be applied which, onfreezing, freezes the specimen 36 to the freezing stage 32. Cooling ofthe freezing stage to freeze the specimen and water, may be effected inknown manner by means of a stream of cold carbon dioxide, or in a novelmanner by means of a number of Peltier cooling junctions built into thefreezing stage 32 and connected to a source of unidirectional current ofappropriate polarity and to .a heat sink having a number of heatingjunctions for dissipating the heat extracted from the freezing stage 32by the Peltier cooling junctions. The arrangement used for cooling thefreezing stage 32 by cooling junctions may be similar to that alreadydescribed with reference to FIGURES 2 and 3 and will not, therefore, beagain described. A thermistor device may also be built into the freezingstage for sensing its temperature and for controlling the currentthrough the cooling junctions and/or providing an indication of thefreezing stage temperature accordingly.

The object clamp holder 34 is mounted on a verticaladjusting mechanism40 which is actuated by a crank handle 41 extending from the undersideof the microtome stand to raise or lower the object clamp holder 34. Amicrometer screw (not shown) accurately controls the movement of thespecimen towards and away from a cutting plane along which the cuttingedge of the knife 12 moves, so that the thickness of the sections takenis always known. To use the microtome the operator slackens the screws11 and fits a microtome knife between the clamping jaws 7 and 8. Thescrews 11 are then tightened so that the metal plates 23 associated withthe cooling junctions 15 are pressed into close contact with theadjacent surfaces of the knife 12. Cooling water is then circulatedbetween the inlet 21 and the outlet 22 of each of the cooling chambers18 and a suitable source of unidirectional current is connected to thetwo terminals 27 with the correct polarity to ensure that the coolingjunctions 15 extract heat and the heating junctions 17 dissipate heat.The temperature of the knife may be determined by the thermistors 24 oralternatively the approach of the temperature of the knife to the valuedesired may be determined from experience of the length of time requiredto cool the knife from room temperature. The knife is cooled "by theextraction of heat via the plates 23, which are cooled by the coolingjunctions 15. The heat extracted by the cooling junctions 15 isliberated by the associated heating junctions 17 and is conducted viathe plates 123 to the cooling fluid flowing through the associatedchambers 18. While the knife is being cooled, the clamp screws 31 can beslackened and the knife set at the desired angle of attack by rotatingthe knife clamp 6 around the carrier arm 4. Also, the specimen to besectioned can be frozen to the freezing stage during this period.

To take sections, the operator grips the knob 5 at the end of thecarrier .arm 4 and swings the arm in a horizontal are around the axis ofthe spindle 3. Return movement of the carrier arm after a cut has takenplace is effected by the spring return and is also arranged to drive thevertical adjusting mechanism 40 to raise the specimen by a thicknessequal to the section which has previously been cut. The thickness of thesection can be varied :by means of a feed setting. Thus, after eachsection is taken, the height of the specimen is automatically adjusted.The microtome may be set for manual adjustment of the specimen heightinstead of the above described automatic adjustment should successivesections of different thickness be desired.

FIGURES 4 and 5 show a knife clamp modified by the replacement of theelectrical cooling units with reservoir cooling units. For the sake ofsimplicity only the pro ection 14 at the left-hand end of the upper jaw7 has been shown in FIGURE 4 and it will be understood that theright-hand projection 14 is similarly constructed. Proection 14 isformed internally with a reservoir 50 which is insulated from thesection 14 by a double layer of thermal insulation 51. The reservoir 50is provided at one end with an inlet port 52 which is normally closed bya closure cap (not shown), and with an outlet port 53 disposed at theother end of the reservoir. The floor of the reservoir is formed by ametal plate having its undersurface constituting a pad 54 ofcomplementary shape to the top surface of the adjacent portion of aknife when gripped between the jaws 7 and 8.

Referring to FIGURE 5 the bottom jaw 8 of the clamp 6 has theknife-engaging surfaces of the two projections 14 covered by a layer ofthermal insulation 55. In this construction of clamp, there is noelectrical control of the knife temperature and thus no thermistors areprovided. All the surfaces of both of the jaws in contact with the knifeapart from the metal pads 54, are covered by a layer of thermalinsulation to reduce the transfer of heat between the clamp and theknife.

To cool a knife, held in the clamp, a cooling fluid, either liquid orgaseous, is circulated through the reservoir 50 between the inlet port52 and the outlet port 53. This coolant fluid may flow around a closedcircuit and by extracting heat continuously from the metal pad 54 theknife temperature is held below a desired maximum value.

The knife clamp shown in FIGURES 4 and 5 may be modified to obtainexceedingly low temperatures of the knife, by plugging up the outletport 53 and admitting liquid air into the reservoir 50 by means of theinlet port 52. Liquid air having a very low temperature rapidly coolsthe knife to a very low temperature. The inlet port should not be closedin order to allow the vaporised liquid air to leave the reservoir 50.

Various modifications of the above described knife clamps are possible.For example, silver pads, which are preferable, may be replaced bycopper pads or aluminium pads to provide a cheaper microtome with verylittle loss of performance.

Although the clamp of the invention has been described with reference toa microtome in which the knife is moved in relation to the specimen tobe sectioned, it will be appreciated that the invention is usable inmicrotomes where the section is moved and the knife is held stationary,and indeed in microtomes where the specimen is mounted between a coupleof guide surfaces, and a knife mounted in a knife clamp adapted to beheld by the hand is moved down the surfaces to section the specimen.

From the above description it will be understood that the inventionprovides an improved microtome knife clamp able to grip a conventionalmicrotome knife and capable of holding the knife at a sufiiciently lowtemperature to prevent any tendency for a frozen section to thaw on theknife and collapse into a shapeless wet lump before it can be examined.

I claim:

1. A microtome comprising, in combination:

(a) a massive stand including a freezing stage holder;

(b) a knife;

(c) means for effecting relative movement between the knife and theholder;

(d) a knife clamp having a pair of jaws for gripping the knife, and amechanism operatively connected to the jaws to open and close them torelease and to hold the knife, respectively;

(e) a heat sink in the jaws, the knife extending into the heat sink whenheld by the jaws;

(f) a thermoelectric cooling unit in the heat sink, said cooling unithaving heating and cooling junctions utilizing the Peltier effect;

(g) heat-conductive pads adjacent the cooling junctions and havingsurfaces in conforming contact with the knife;

(h) heat transmitting elements adjacent the heating junctions and incontact with the heat sink; and

(i) electrical circuit means extending through the jaws to the heatingand cooling junctions.

2. The microtome of claim 1, wherein said junctions are bondedrespectively to said pad and to said heat transmittin g element.

3. The microtome of claim 1, wherein said jaws define cooling chambersadjacent the heat transmitting elements, the jaws having inlet andoutlet ports communicating with the chambers to circulate a collantfluid therethrough.

4. The microtome of claim 1, further comprising a thermistor bonded tothe thermoelectric cooling unit adjacent the knife cutting edge, theelectrical circuit means being connected also to the thermistor.

5. The microtome of claim 1, further comprising a layer of heatinsulating material between all surfaces of the jaws and the knife atlocations where the knife is not in contact with the heat-conductivepads.

(References on following page) References Cited by the Examiner UNITEDSTATES PATENTS Rusterholz 83700 Mac Donald 83699 Ott 83-699 Turner83915.5 Jacoby 83 915.5 Herbain 83-915.5

Sparer 83-915.5 Tuck 136-4.2 Baude 323-69 Hirschhorn 62--3 Meyers 83171ANDREW R. JUHASZ, Primary Examiner.

WILLIAM W. DYER, JR., Examiner.

1. A MICROTOME COMPRISING, IN COMBINATION: (A) A MASSIVE STAND INCLUDINGA FREEZING STAGE HOLDER; (B) A KNIFE; (C) MEANS FOR EFFECTING RELATIVEMOVEMENT BETWEEN THE KNIFE AND THE HOLDER; (D) A KNIFE CLAMP HAVING APAIR OF JAWS FOR GRIPPING THE KNIFE, AND A MECHANISM OPERATIVELYCONNECTED TO THE JAWS TO OPEN AND CLSE THEM TO RELEASE AND TO HOLD THEKNIFE, RESPECTIVELY; (E) A HEAT SINK IN THE JAWS, THE KNIFE EXTENDINGINTO THE HEAT SINK WHEN HELD Y THE JAWS; (F) A THERMOELECTRIC COOLINGUNIT IN THE HEAT SINK, SAID COOLING UNIT HAVING HEATING AND COOLINGJUNCTIONS UTILIZING THE PELTIER EFFECT; (G) HEAT-CONDUCTIVE PADSADJACENT THE COOLING JUNCTIONS AND HAVING SURFACES IN CONFORMING CONTACTWITH THE KNIFE; (H) HEAT TRANSMITTING ELEMENTS ADJACENT THE HEATINGJUNCTIONS AND IN CONTACT WITH THE HEAT SINK; AND (I) ELECTRICAL CIRCUITMEANS EXTENDING THROUGH THE JAWS TO THE HEATING AND COOLING JUNCTIONS.